The newly available images of ribosomes at the atomic scale mark a critical milestone in the quest to link ribosome structure with function. One important function of the ribosome is to faithfully maintain translational reading frame. Viral mRNA signals that abrogate this function by programming ribosomes to shift frame have proved to be of useful in elucidating the molecular mechanisms underlying this essential task. Our studies on "Programmed Ribosomal Frameshifting" (PRF) have begun to link ribosome structure with this aspect of its function. We have shown that both the biophysical interactions between ribosomal proteins, rRNAs and tRNAs, and the biochemical properties of ribosome-associated enzymatic activities are important for proper reading frame maintenance. Structural analyses of mutants of ribosomal proteins and rRNAs were previously identified as affecting PRF reveals a central commonality: all of the phenotype producing mutations occur where these proteins/rRNAs physically interact with important regions of other components of the large ribosomal subunit. Based on these findings, we )ropose to use a targeted mutagenesis approach in an effort to further define how ribosome structure influences function. Specifically, we will: (I) determine the effects of targeted mutations of specific ribosomal proteins on yeast ribosome structure and function; (11)determine the effects of targeted mutations of specific bases in the yeast 25S rRNA on yeast ribosome structure and function, and (111) determine the effects of mutants having rRNA modification defects on ribosome structure and function. Concurrent to this work, we anticipate that other researchers will elucidate the structures of ribosomes from yeast and other eukaryotes at the atomic scale. Thus, the proposed studies will uniquely position us to address questions relating to eukaryotic ribosome function at a point in history when the atomic level structure is being revealed. By the end of the grant period, we anticipate having made a significant contribution to the general understanding of how ribosome structure influences function.